Abstract
The calpain activity in cells can be experimentally manipulated in vitro by calpain inhibitors, and various types of calpain inhibitors such as peptide aldehydes and α-mercapto-acrylic acid derivatives are widely used as a valuable tool to elucidate the physiological and pathological roles of calpain. Here I describe the experimental procedures with calpain inhibitors, with human neutrophils being primarily used in this experiment. It should be noted that potent calpain inhibitors not only inhibit the calpain activity but also stimulate cell functions via direct activation of human formyl peptide receptors and/or other G protein-coupled receptors depending on the inhibitors used.
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References
Lokuta MA, Nuzzi PA, Huttenlocher A (2003) Calpain regulates neutrophil chemotaxis. Proc Natl Acad Sci U S A 100:4006–4011
Katsube M, Kato T, Kitagawa M, Noma H, Fujita H, Kitagawa S (2008) Calpain-mediated regulation of the distinct signaling pathways and cell migration in human neutrophils. J Leukoc Biol 84:255–263
Ozaki Y, Kato T, Kitagawa M, Fujita H, Kitagawa S (2008) Calpain inhibition delays neutrophil apoptosis via cyclic AMP-independent activation of protein kinase A and protein kinase A-mediated stabilization of Mcl-1 and X-linked inhibitor of apoptosis (XIAP). Arch Biochem Biophys 477:227–231
Camins A, Verdaguer E, Folch J, Pallas M (2006) Involvement of calpain activation in neurodegenerative processes. CNS Drug Rev 12:135–148
Mani SK, Balasubramanian S, Zavadzkas JA, Jeffords LB, Rivers WT, Zile MR, Mukherjee R, Spinale FG, Kuppuswamy D (2009) Calpain inhibition preserves myocardial structure and function following myocardial infarction. Am J Physiol Heart Circ Physiol 297:H1744–H1751
Noma H, Kato T, Fujita H, Kitagawa M, Yamano T, Kitagawa S (2009) Calpain inhibition induces activation of the distinct signalling pathways and cell migration in human monocytes. Immunology 128:e487–e496
Sasaki T, Kishi M, Saito M, Tanaka T, Higuchi N, Kominam E, Katunuma N, Murachi T (1990) Inhibitory effect of di- and tripeptidyl aldehydes on calpains and cathepsins. J Enzym Inhib 3:195–201
Wang KK, Nath R, Posner A, Raser KJ, Buroker-Kilgore M, Hajimohammadreza I, Probert AW Jr, Marcoux FW, Ye Q, Takano E, Hatanaka M, Maki M, Caner H, Collins JL, Fergus A, Lee KS, Lunney EA, Hays SJ, Yuen P (1996) An alpha-mercaptoacrylic acid derivative is a selective nonpeptide cell-permeable calpain inhibitor and is neuroprotective. Proc Natl Acad Sci U S A 93:6687–6692
Rock KL, Gramm C, Rothstein L, Clark K, Stein R, Dick L, Hwang D, Goldberg AL (1994) Inhibitors of the proteasome block the degradation of most cell proteins and the generation of peptides presented on MHC class I molecules. Cell 78:761–771
Fujita H, Kato T, Watanabe N, Takahashi T, Kitagawa S (2011) Calpain inhibitors stimulate phagocyte functions via activation of human formyl peptide receptors. Arch Biochem Biophys 513:51–60
Fujita H, Kato T, Watanabe N, Takahashi T, Kitagawa S (2011) Stimulation of human formyl peptide receptors by calpain inhibitors: homology modeling of receptors and ligand docking simulation. Arch Biochem Biophys 516:121–127
Kitagawa S, Kato T, Kitagawa M, Aomatsu M, Fujita H (2013) Biological effects of calpain inhibitors on human phagocyte functions. Chapter V. In: Lashinski EM (ed) Enzymes and enzyme activity. Nova Science Publishers, New York, pp 121–144
Kobayashi S, Yamashita K, Takeoka T, Ohtsuki T, Suzuki Y, Takahashi R, Yamamoto K, Kaufmann SH, Uchiyama T, Sasada M, Takahashi A (2002) Calpain-mediated X-linked inhibitor of apoptosis degradation in neutrophil apoptosis and its impairment in chronic neutrophilic leukemia. J Biol Chem 277:33968–33977
Derouet M, Thomas L, Cross A, Moots RJ, Edwards SW (2004) Granulocyte macrophage colony-stimulating factor signaling and proteasome inhibition delay neutrophil apoptosis by increasing the stability of Mcl-1. J Biol Chem 279:26915–26921
Kato T, Kutsuna H, Oshitani N, Kitagawa S (2006) Cyclic AMP delays neutrophil apoptosis via stabilization of Mcl-1. FEBS Lett 580:4582–4458
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Kitagawa, S. (2019). Experimental Manipulation of Calpain Activity In Vitro. In: Messer, J. (eds) Calpain. Methods in Molecular Biology, vol 1915. Springer, New York, NY. https://doi.org/10.1007/978-1-4939-8988-1_16
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DOI: https://doi.org/10.1007/978-1-4939-8988-1_16
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